Members of the tumor-necrosis factor (TNF) family of cytokines are involved in an ever expanding array of critical biological functions. Each member of the TNF family acts by binding to one or more members of a parallel family of receptor proteins. These receptors in turn signal intracellularly to induce a wide range of physiological or developmental responses. Many of the receptor signals influence cell fate, and often trigger terminal differentiation. Examples of cellular differentiation include proliferation, maturation, migration, and death.
TNF family members are Type II membrane bound proteins, having a short intracellular N-terminal domain, a transmembrane domain, and the C-terminal receptor binding domains lying outside the cell surface. In some cases the extracellular portion of the protein is cleaved off, creating a secreted form of the cytokine. While the membrane bound proteins act locally, presumably through cell contact mediated interaction with their receptors, the secreted forms have the potential to circulate or diffuse, and therefore can act at distant sites. Both membrane bound and secreted forms exist as trimers, and are thought to transduce their signal to receptors by facilitating receptor clustering.
The TNF receptor protein family is characterized by having one or more cysteine rich extracellular domains. Each cysteine rich region creates a disulfide-bonded core domain, which contributes to the three dimensional structure that forms the ligand binding pocket. The receptors are Type I membrane bound proteins, in which the extracellular domain is encoded by the N-terminus, followed by a transmembrane domain and a C-terminal intracellular domain. The intracellular domain is responsible for receptor signaling. Some receptors contain an intracellular “death domain”, which can signal cell apoptosis, and these can be strong inducers of cell death. Another class of receptors can weakly induce cell death; these appear to lack a death domain. A third class of receptors do not induce cell death. All classes of receptors can signal cell proliferation or differentiation instead of death, depending on cell type or the occurrence of other signals.
A well studied example of the pluripotent nature of TNF family activity is the nominant member, TNF. TNF can exist as a membrane bound cytokine or can be cleaved and secreted. Both forms bind to the two TNF receptors, TNF-R55 and TNF-R75. Originally described on the basis on its' ability to directly kill tumor cells, TNF also controls a wide array of immune processes, including inducing acute inflammatory reactions, as well as maintaining lymphoid tissue homeostasis. Because of the dual role this cytokine can play in various pathological settings, both agonist and antagonist reagents have been developed as modifiers of disease. For example TNF and LTα (which also signals through the TNF receptors) have been used in treatment for cancers, especially those residing in peripheral sites, such as limb sarcomas. In this setting direct signaling by the cytokine through the receptor induces tumor cell death (Aggarwal and Natarajan, 1996. Eur Cytokine Netw 7:93-124).
In immunological settings, agents that block TNF receptor signaling (e.g., anti-TNF mAb, soluble TNF-R fusion proteins) have been used to treat diseases like rheumatoid arthritis and inflammatory bowel disease. In these pathologies TNF acts to induce cell proliferation and effector function, thereby exacerbating autoimmune disease, and in this setting blocking TNF binding to its receptor(s) has therapeutic benefit (Beutler, 1999. J Rheumatol 26 Suppl 57:16-21).
A more recently discovered ligand/receptor system appears amenable to similar manipulations. Lymphotoxin beta (LTβ), a TNF family member which forms heterotrimers with LTα, bind to the LTβ-R. Some adenocarcinoma tumor cells which express LTβ-R can be killed or differentiated when treated with an agonistic anti-LTβ-R mAb (Browning et al., 1996. J Exp Med 183: 867-878). In immunological settings it has been shown that anti-LTβ mAb or soluble LTβ-R-Ig fusion protein can block the development of inflammatory bowel diseases, possibly by influencing dendritic cell and T cell interaction (Mackay et al., 1998. Gastroenterology 115:1464-1475).
The TRAIL system also has potential as a cancer therapy. TRAIL interacts with a number of membrane bound and soluble receptors. Two of these receptors, TRAIL-R1 and TRAIL R2 (also called DR4 and DR5), transmit death inducing signals to tumor cells but not to normal cells, which express additional TRAIL receptors that do not induce death. These additional receptors are thought to function as decoys. The use of soluble TRAIL to kill tumor cells relies on the selective expression of decoy receptors on normal but tumor tissue (Gura, 1997. Science 277: 768).
Tumor cells themselves often express a variety of decoy receptors that block immune recognition or effector functions. Indeed some tumors overexpress TRAIL decoy receptors, apparently to avoid TRAIL mediated death (Sheikh et al., 1999. Oncogene 18: 4153-4159). This limits the utility of TRAIL as an anti-tumor agent in some settings. Similar observations have been made about a decoy receptor for FAS-L, which is overexpressed by lung and colon cancer cells (Pitti et al., 1998. Nature 396: 699-703), and for the IL-1 receptor antagonist (Mantovani et al., 1998. Ann. N Y Acad. Sci. 840: 338-351). Decoy receptors are also employed by viral genomes to protect infected host cells from host defense mechanisms.
APRIL (A Proliferation Inducing Ligand) is a new member of the TNF family of proteins. APRIL expression and functional studies suggest that this protein is utilized by tumor cells to induce rapid proliferation. Tumor cell lines treated with soluble APRIL protein or transfected with APRIL cDNA grow rapidly in vitro. APRIL transfected cells implanted into immunodeficient mice grow rapidly as tumors. Finally, human tumor cells, but not normal tissue, express high levels of APRIL messenger RNA. These observations suggest that APRIL binds to a receptor that is also expressed by tumor cells, setting up autocrine or paracrine tumor cell activation. In addition, it is possible that APRIL acts in other disease settings, such that activating or blocking the APRIL pathway would have additional utility. For example, underexpression or overexpression of APRIL may play a role in developmental defects, since development is often characterized by the carefully controlled balance between cell proliferation and cell death. Similarly, APRIL may act in cell proliferative diseases, such as those that occur in connection with some autoimmune diseases (e.g., lupus) or in inflammatory diseases where cell populations expand rapidly (e.g., bacterial sepsis).
Based on the known utility of using agonists and antagonists of TNF and TNF receptor family members as disease modifiers, the APRIL pathway presents itself as an important target for drug development. This is particularly true for cancer therapy since tumor cells appear to produce and utilize APRIL to support their own growth, and are therefore unlikely to produce decoy receptors or other antagonists of the APRIL pathway. Thus the APRIL pathway is uniquely different from, for example, the TRAIL or FAS-L pathways, which can be thwarted by tumor decoy receptors.
Current treatments for cancer are inadequate for many tumor types, due to poor efficacy, low impact on survivorship, toxicity that causes severe side effects, or combinations thereof. Therefore there is a need to identify and develop additional methods for treating cancer growth which can provide efficacy without inducing severe side effects. Antagonists of the APRIL pathway, including anti-APRIL mAbs, anti-APRIL receptor mAbs, soluble APRIL receptor-Ig fusion proteins, natural antagonists, small molecule antagonists, and chemical, pharmaceutical, or other antagonists would thus be useful.
To this end we have identified B cell mediated protein (BCM or BCMA) as a receptor for APRIL.